It is the objective of the proposed research to relate alterations in the level of cellular polyunsaturated fatty acids (PUFA) to differences in the longevity of WI-38 cells in vitro, and to correlate these differences with the susceptibility of cell lipids to peroxidation. To accomplish this, cells will be grown in delipidized serum in the presence of exogenous linolenic or oleic acid. The toxicity observed when fatty acid salts are added in discrete doses to cells in culture will be obviated by using an infusion pump to slowly and continuously add dilute solutions of the fatty acids over a period of two generations to cultures in log phase. This system has been effectively utilized with mouse L-fibroblasts to increase their PUFA content from 5% to 50% without causing any change in growth rate. Two protocols will be employed in the proposed research. In the first, cells at different passage levels will be infused as described and then exposed to high dose rates of ionizing radiation. Plating efficiencies will be compared to those of non-infused control cells of the same age, and related to the PUFA content and the amount of lipid peroxidation occurring after each treatment. In the second protocol, cells throughout their in vitro life span will be alternately infused and exposed to ionizing radiation at low dose rates. The longevity of the culture will be correlated with PUFA content and the amount of lipid peroxidation occurring in the cells. The role of tocopherol in increasing the life span of cells will be further investigated by determining whether cells with high PUFA content and cells with low PUFA content are protected, to the same extent, from aging and lipid peroxidation. The results of these experiments should be useful in elucidating the role of PUFA, tocopherol and lipid peroxidation in the in vitro aging process.